JP2002360665A - Soft capsule for improving disintegration property - Google Patents

Abstract

PROBLEM TO BE SOLVED: To provide a soft capsule for improving a disintegration property. SOLUTION: This soft capsule of a multiplayer structure has the innermost layer and one or two or more coating film layers successively laminated so as to cover the innermost layer, and is constituted so that at least one coating film layer of the coating film layers includes an enzyme, and decomposes the coating film layer including the enzyme or the other coating film layer. A capsule of a three-layer structure having the innermost layer, an inner coating film layer, and a dry outer coating film layer is desirable. The enzyme for decomposing the inner coating film is included in the outer coating film layer, and an inner coating film is disintegrated when the enzyme acts by supplying moisture.

Description

DETAILED DESCRIPTION OF THE INVENTION

[0001]

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a soft capsule which has a high storage stability in a dry state and, when it is transferred to a state with moisture, a film layer collapses in a desired time and the inside and outside of the capsule are released. More specifically, the present invention relates to a soft capsule having a structure of two or more layers, wherein the capsule contains an enzyme for decomposing the film layer or an adjacent film layer.

[0002]

2. Description of the Related Art With the recent development of technology, liquids, powders,
It has become possible to encapsulate cells and the like. However, it is still difficult to quickly disintegrate the capsule under the desired conditions. In particular, disintegration of capsules and release of drugs at desired sites such as the intestine and large intestine in so-called drug delivery systems, release of encapsulated pesticides sprayed on soil under appropriate moisture conditions, or seeding on soil or growing containers Although it is desired to induce germination of artificial seeds by rapid force collapse, capsules having satisfactory desired disintegration properties have not yet been provided.

[0003]

Therefore, a desired part,
There is a need for capsules that disintegrate depending on location, conditions, and the like. If such a capsule is obtained, various pharmaceuticals exhibiting activity at the desired site, transport of useful microorganisms such as lactic acid bacteria to the desired site, and further applied to tissues such as somatic embryos to produce artificial seeds A wide range of applications are possible.

[0004]

Means for Solving the Problems The present inventors have conducted intensive studies in order to solve the above-mentioned problems, and as a result, the dried capsule containing an enzyme or a microorganism capable of decomposing the film layer in the film layer of the capsule. Succeeded in obtaining a capsule, placing the obtained capsule in the desired location and controlling the moisture and temperature, the enzymes or microorganisms contained began to work, and it was recognized that the film collapsed at a rate according to the conditions, The present invention has been completed.

That is, the present invention provides a soft capsule having a multilayer structure having an innermost layer and one or more coating layers sequentially laminated so as to cover the innermost layer, wherein at least one of the coating layers is an enzyme. Alternatively, the present invention relates to a soft capsule containing a microorganism and configured to decompose a film layer or another film layer containing the enzyme or the microorganism under a predetermined condition.

[0006] In a preferred embodiment, the enzyme or the microorganism is contained in the outermost coating layer of the soft capsule, and the outermost coating layer is dried.

In a more preferred embodiment, the soft capsule is a three-layered capsule having an innermost layer, an inner coating layer covering the innermost layer, and an outer coating layer covering the inner coating layer. The coating layer contains enzymes or microorganisms capable of decomposing the inner coating layer.

[0008] In a preferred embodiment, the coating layer of the dried soft capsule is a coating layer formed of a substance selected from the group consisting of proteins, polysaccharides, hardened fats and oils, and biodegradable plastics.

[0009] In a preferred embodiment, the enzyme is a protease, a polysaccharide-degrading enzyme, a hydrogenated fat-degrading enzyme, or an esterase.

In a further preferred embodiment, the innermost layer contains a medicament, a pesticide or a regenerable plant cell tissue, and the inner coat layer is mainly composed of hardened fat and oil,
The outer coat layer is a dried outer coat layer containing an enzyme or a microorganism capable of decomposing the hardened oil and fat.

[0011] In another preferred embodiment, the soft capsule is selected from the group consisting of a seamless soft capsule and a seamed soft capsule.

Another aspect of the present invention is a three-layer structure comprising an innermost layer containing a regenerable plant cell tissue, an inner coat layer covering the innermost layer, and an outer coat layer covering the inner coat layer. Wherein the inner shell layer is a hardened oil and fat as a main component, and the outer shell layer is a dried outer shell layer containing an enzyme or a microorganism capable of decomposing the hardened oil and fat.

[0013]

DESCRIPTION OF THE PREFERRED EMBODIMENTS (Structure of Soft Capsule of the Present Invention) The soft capsule of the present invention is a soft capsule having a multilayer structure having an innermost layer and one or more coating layers sequentially laminated so as to cover the innermost layer. . Therefore, it may have a two-layer structure, a three-layer structure, a four-layer structure, or the like. For example, in the case of a three-layer structure, the structure shown in FIG. 1 can be taken. FIG.
FIG. 2 is a schematic diagram when a soft capsule is used as an artificial seed.

In the case of a two-layer structure, it is composed of an innermost layer and a coating layer (outermost layer) covering the innermost layer. The film layer (outermost layer) contains an enzyme or a microorganism capable of decomposing the film layer. This coating layer (outermost layer) is preferably dried for the purpose of suppressing the enzyme activity.

In the case of the three-layer structure, as shown in FIG. 1, it is composed of an innermost layer, an inner coating layer covering the innermost layer, and an outer coating layer covering the inner coating layer. At least one of the inner coating layer and the outer coating layer contains an enzyme or a microorganism capable of decomposing any of the coating layers (hereinafter, collectively referred to as an enzyme or the like). For example, the inner coat layer may contain an enzyme capable of decomposing the inner coat layer, an enzyme capable of decomposing the outer coat layer, or both enzymes. The outer coat layer may contain an enzyme capable of decomposing the outer coat layer, an enzyme capable of decomposing the inner coat layer, or both enzymes. When a film layer containing an enzyme or the like is decomposed by the enzyme or the like, the film layer is preferably dried to suppress the enzyme activity.

In a preferred soft capsule having a three-layer structure, the outer coat layer contains an enzyme capable of decomposing the inner coat layer, but the inner coat layer does not contain an enzyme or the like. Since it is preferable to suppress the enzyme activity in the outer coat layer from the viewpoint of preventing the decomposition of the inner coat layer during storage, the outer coat layer is preferably dried.

In the soft capsule of the present invention, the polarities of adjacent layers are preferably different from each other. For example, the layers are preferably alternately different in polarity, such as a hydrophilic layer / oil layer / hydrophilic layer. . When the innermost layer is an aqueous material, it is important that the coating layer covering the innermost layer is a water-resistant oily layer, not only for forming the capsule, but also for preventing the evaporation of moisture in the innermost layer. It is.

(Innermost layer) The innermost layer of the capsule contains a substance which is released under a desired place and condition. The material contained in the innermost layer can be hydrophilic, oily, or powder.
Substances that can be contained in the innermost layer include pharmaceuticals, agricultural chemicals, microbial cells, regenerable plant cell tissues, and the like.
The medicine contained in the innermost layer is not particularly limited. For example, in the case of crystalline powder such as cilostazol having an antiplatelet effect, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) are suspended in hardened oil and fat.
The fats and oils such as can be contained in the innermost layer in the form as it is or in the form of an emulsion. Examples of microbial cells include useful microorganisms such as bifidobacteria and lactic acid bacteria. These microorganisms are contained in the innermost layer as an aqueous solution or as a dry powder. In the case of a dry powder, it can be used by suspending it in a liquid fat or oil. As the regenerated plant cell tissue, adventitious embryos, adventitious buds, polyblasts, shoot tips, growing points, adventitious roots, hairy roots, and protocol-like spheres are preferably used. Virus-free tissues can also be used.

(Coating layer) A coating layer (first layer) adjacent to the innermost layer
When the innermost layer is hydrophilic, it is preferable that the innermost layer be composed of a lipophilic substance. As such a lipophilic substance, hardened fats and oils are preferably used. The next coating layer (second coating layer) is a layer made of a hydrophilic substance, and the next coating layer (third coating layer) is preferably made of an oily substance.

When the innermost layer is made of a lipophilic substance, the interface of the coating layer (first coating layer) adjacent to the innermost layer is not clear, but it may be composed of a lipophilic substance, May be constituted.

When the first coating layer is a lipophilic substance,
The fat in the innermost layer may be a hardened fat which is solid at normal temperature and has a different melting point and which is decomposed by an enzyme. In this case, the next coating layer (second coating layer) is a layer made of a hydrophilic substance,
The next coating layer (third coating layer) is preferably made of an oily substance.

When the first coating layer is a hydrophilic substance,
It must be a substance that can be decomposed by enzymes and the like. Such substances include proteins, polysaccharides, biodegradable bioplastics, and the like. These can be used alone or in combination of two or more. The next coating layer (second coating layer) is preferably a layer made of a lipophilic substance, and the next coating layer (third coating layer) is preferably made of a hydrophilic substance.

(Outermost Layer) The outermost coating layer is determined by the type of innermost layer and the number of layers. When the innermost layer is hydrophilic, if the capsule has a two-layer structure, the outermost layer is a lipophilic film layer, and if the capsule has a three-layer structure, it is a hydrophilic film layer. The outermost layer is preferably dried from the viewpoint of suppressing the enzyme activity.

(Material of film layer) The film layer may be any material as long as it can be decomposed by an enzyme or the like. Therefore, it is not necessary that the entire coating layer is decomposed by the substrate of these enzymes or microorganisms, but is used as a part of the coating layer and contained to such an extent that the coating layer can be disintegrated by being decomposed. It may be.

As the lipophilic substance among the substances used for the coating layer, hardened fats and oils are preferably used. The hardened oils and fats include oils and fats containing hardened oil as a main component, and include oils and fats of hardened oil alone or oils and fats prepared by mixing hardened oil with other oils and fats to have desired properties. Preferably, it is solid at room temperature, has a melting point according to the purpose, and is hydrolyzed by lipases. As hardened fats and oils,
Examples thereof include medium chain fatty acid triglycerides and diglycerides, and examples thereof include butter, margarine, shortening and cocoa butter, but are not limited thereto.

As the hydrophilic substance, proteins, polysaccharides and biodegradable plastics are preferably used.

Examples of the protein include, but are not limited to, gelatin and collagen. These can be used alone or in combination of two or more. Particularly preferred proteins include, after drying,
Gelatin having a high oxygen barrier property is exemplified.

As the polysaccharide, a gel-forming polysaccharide is preferably used. Such polysaccharides include starch, amylose, polygalacturonic acid, agar, carrageenan, gum arabic, gellan gum, xanthan gum,
Examples include, but are not limited to, pectin, alginic acid, and the like. These can be used alone or in combination of two or more.

Examples of biodegradable plastics include, but are not limited to, polylactic acid, polyhydroxybutyric acid, polyglutamic acid, and mixtures thereof. These can be used alone or in combination of two or more.

Further, if necessary, the properties of the coating layer can be changed by adding sugar, sugar alcohol, polyhydric alcohol, pullulan, chitosan, chitin, amino acid and the like to the biodegradable substance. .

(Enzymes or microorganisms that decompose the film layer) Examples of enzymes that decompose hydrogenated oils and fats include lipases and microorganisms that can produce lipase. Lipase producing microorganisms include Aspergillus niger (Asperg
illus niger, Candida rug
osa). When used as pesticides or artificial seeds, lipase for detergents can be used, and when capsules are used for pharmaceuticals and foods, lipases for food processing can be used. The amount of the enzyme or the microorganism to be added varies depending on the purpose of use and the conditions of use of the capsule.

Examples of the enzymes that degrade proteins include proteases and microorganisms that produce proteases. When an enzyme is used, it may be necessary to select proteases (proteolytic enzymes) according to the type of protein used.
Examples of microorganisms that produce protease include Bacillus subtilis.

Examples of the enzyme for decomposing the polysaccharide include enzymes and microorganisms according to the kind of the polysaccharide used.
For example, in the case of starch, amylases are used as enzymes, and the microorganisms producing amylase include Bacillus subtilis. In the case of polygalacturonic acid, polygalacturonase is used, and as a microorganism that produces polygalacturonase, Aspergillus niger (Aspergillus niger) is used.
niger). In the case of pectin, pectinase is used, and as a microorganism producing pectinase, Aspergillus niger and the like can be mentioned.

Polylactic acid which is a biodegradable plastic,
Examples of enzymes that degrade polyhydroxybutyric acid include esterases, and microorganisms that produce these esterases include microorganisms of the genus Rhizopus. Examples of enzymes that degrade polyglutamic acid include peptidases, and examples of microorganisms that produce peptidase include Aspergillus oryzae.

The above examples relating to the combination of the biodegradable substance used for the coating layer of the capsule of the present invention and the enzyme used for decomposing the biodegradable substance or the microorganism producing the enzyme are only examples. The present invention is not limited to the present invention, and a person skilled in the art can appropriately select a combination of a biodegradable substance and an enzyme or a microorganism capable of decomposing the biodegradable substance from commonly used books such as enzymology and biochemistry.

(Method for Producing the Soft Capsule of the Present Invention) The soft capsule of the present invention is produced by a method such as a rotary method or a dropping method usually used for producing by a person skilled in the art. The soft capsule obtained by a technique well-known to those skilled in the art can be a seamless soft capsule, a seamless, so-called seamless soft capsule, preferably,
After molding, the outermost layer is dried.

Hereinafter, a case where a three-layer seamless soft capsule comprising an innermost layer, a hardened oil / fat layer (inner coating layer), and a gelatin layer (outer coating layer) will be specifically described. It is most preferable that the seamless soft capsule is manufactured by a dropping method using a triple nozzle. A method for producing a seamless capsule is described in, for example, Bioscience and Industry, vol. 58, No. 7, p.
0). The basic production method of this seamless soft capsule is a method already used industrially, and therefore, mass production is easy.

First, the target substance in the innermost layer is prepared by dispersing or dissolving in a liquid oil or fat or an aqueous solution. The hardened fat or oil forming the inner coat layer is dissolved, and on the other hand, an appropriate amount of lipase is added to a gelatin solution of an appropriate concentration (for example, 18%) to prepare a gelatin solution for forming the outer coat layer.
In a triple tube nozzle, put the internal solution, hardened oil and fat, and gelatin solution, using a seamless soft capsule manufacturing machine,
Encapsulation is performed by the dropping method in liquid. Capsule droplet formation and solidification are carried out in coagulated oil at a temperature as low as possible in order to suppress the enzymatic activity, to give a three-layered capsule. After removing the coagulated oil from the obtained capsule, the dried capsule is quickly dried to obtain a dried capsule. Drying is preferably performed promptly at a low temperature so that lipase does not work during drying.

When a rugby ball-type soft capsule is obtained by a rotary capsule manufacturing method, it has a two-layer structure of an innermost layer and a coating layer (outermost layer). When gelatin is used for the film layer (outermost layer), a dried gelatin sheet containing a protease or a microorganism is used.

Alternatively, a soft capsule is produced without adding a protease to a gelatin solution or a gelatin film for forming a film layer (skin layer), and then the protease is dissolved on a sugar solution or the like to coat the surface. Then, it may be dried promptly. In the case of dissolving or dispersing the target substance in hardened fat or oil as a content, the surface of the gelatin film of the dried soft capsule containing lipase is dissolved and coated with proteolytic enzyme in a sugar solution, etc. May be dried.

The obtained soft capsule can be sufficiently oxidized by an enzyme or a microorganism under a predetermined condition, for example, when water is supplied, when an appropriate pH is set, or when an appropriate temperature is set. When placed in an environment where it can perform, the film layer containing the enzyme or microorganism or the adjacent film layer is degraded.

The soft capsules of the present invention thus obtained are completely different from the single-sphere beads of calcium alginate gel described as capsules in the descriptions of various scientific and patent documents. Furthermore, they are different from wet calcium alginate gel beads.

(Enteric Bifidobacterium Capsules) Enteric bifidobacteria capsules are prepared by suspending a dried bifidobacterium powder in a heated and melted hardened oil and fat, and using this as the innermost layer. A seamless soft capsule with a three-layer structure consisting of a solution in suspension-hardened fat / oil, hardened fat / oil layer, and acid-resistant coating layer (protein-degrading enzyme and pectin-containing gelatin layer), which is quickly dried at low temperature to dry the outermost protein layer To be obtained. The resulting enteric-coated Bifidobacterium capsule absorbs moisture, whereby the outermost layer of the protein is degraded and the hardened oil / fat layer disintegrates under the action of lipase in the intestine, releasing the Bifidobacterium into the intestine .

(Artificial seed) Cell tissue (adventitious embryo, protocomb-like body (hereinafter referred to as
PLB), etc.) and an appropriately sized somatic embryo or PLB
Or the like, and a regenerated plant cell tissue is obtained, suspended in a medium, and used as the contents of the capsule (the innermost layer). A hardened fat or oil which is solid at room temperature is used as the inner film layer, and the outer film layer is prepared as a solution (for example, a 22% gelatin solution) of the biodegradable substance having an appropriate concentration.
(00 units / g), and encapsulated by a submerged dropping method using a triple tube nozzle seamless soft capsule manufacturing machine. At this time, a pump is prepared so that the adventitious embryos of the contents enter one by one. By performing the same treatment as above, an encapsulated artificial seed having the outermost outer coating layer dried is obtained. Drying is preferably performed promptly at a low temperature so that cell tissues do not die and lipase does not work. FIG. 1 shows a schematic view of the artificial seed thus obtained.

The amount of the lipase to be added to the outer coat varies depending on the specific activity of the lipase, but when the artificial seed is sown in soil or a seedling raising pot and watered, the inner coat layer is hardened under the temperature conditions. The amount is such that the fat or oil is decomposed in 6 hours to 14 days. More preferably, the amount is such that the activity is decomposed in one to seven days.

The particle size of the obtained encapsulated artificial seed varies depending on the size of the cell tissue, but is 1 mm to 12 mm.
It is. More preferably, it is 3 mm to 10 mm.

The thus-obtained artificial seed of the present invention can be stored for more than two months while maintaining its germination ability at room temperature in a dry barn or the like, and can be stored especially in a refrigerator or low-temperature water. No need to save. If stored in a refrigerator at 10 ° C or lower, long-term storage of 4 months or more is possible.

The encapsulated artificial seed of the present invention is sown in the soil and sprayed with water to cause the gelatin layer to swell if the outer coat layer is gelatin, followed by the lipase Decomposes rapidly by the action of
At that time, it is considered that oxygen permeability increases, awakens from a dormant state, and shifts to a germinating state.

(Plant Growth Hormone) In the case of plant growth hormone, the aqueous solution of plant growth hormone is used as the innermost layer, the inner film layer is made of hardened oil and fat, and the outer film layer is made of gelatin mixed with lipase. Get a capsule. By embedding this capsule in the root of the crop, the hormone does not act in a dry state, but the capsule disintegrates quickly due to watering or rainfall, and the hormone acts.

[0050]

EXAMPLES Hereinafter, the present invention will be described more specifically with reference to examples, but it goes without saying that the present invention is not limited to these examples.

(Example 1: Dye-encapsulated capsule) Seamless three-layer drying using edible red No. 2 aqueous solution as the content liquid (innermost layer), hardened oil and fat as inner coating layer, and lipase-added gelatin as outer coating layer Capsules were prepared.

More specifically, 0.1% (w /
v) The aqueous solution is used as the content liquid (innermost layer), and the inner coating layer has a melting point of 3
Hardened oil of triglyceride at 2 ° C (Pharmazol B-11)
5, manufactured by Nippon Yushi Co., Ltd.) and 22%
(W / v) Liquid lipase (Novozyme 3)
98, Novo Nordisk Bioindustry Co., Ltd.
) Were mixed at various concentrations (w / v) as shown in Table 1. The volume ratio of each layer is as follows: content liquid layer / inner coating layer /
Outer coating layer = 3: 3: 4. Encapsulation was carried out in a conventional manner using a triple tube nozzle aseptic seamless soft capsule manufacturing machine in a solidified oil, and immediately after deoiling, the drum was dried while sending dry air at 10 ° C.
The particle size of the surface-dried three-layer capsule was 7 mm.

These capsules were stable after being stored in a polyethylene bag containing silica gel and stored at 20 ° C. for 3 months without the red color of the contents appearing on the surface.

The capsules immediately after preparation and the capsules stored for 3 months under the above conditions were placed in a Petri dish with 50 tablets each, sprayed with water uniformly by spraying, and kept warm in a 25 ° C. incubator. The time during which red spots of the content liquid were observed on the surface was observed over time, and 50 particles were averaged to determine the film disintegration time. Table 1 shows the results.

[0055]

[Table 1]

As shown in Table 1, in the capsule containing no lipase in the outer coat layer, the time required for the collapse of the inner coat layer was extremely long. On the other hand, in the capsule containing the lipase in the outer film layer, the film disintegration time was remarkably shortened, and as the amount of lipase added increased, the time required for decomposing the hardened oil and fat as the inner film layer became shorter. Even after storage for 3 months, no evaporation of the water content of the content liquid wrapped with the hardened fat and gelatin was observed. It was observed that the lipase activity was reduced by about 8%, but the effect of the lipase addition was almost the same. From this, it was shown that the capsule of the present invention was extremely stable in a dry state even at room temperature storage, and could withstand long-term storage. Furthermore, it was shown that by adjusting the amount of lipase added to the outer coat layer, it is possible to supply water and to disintegrate the coat layer in a desired time.

Example 2 Artificial Seed Encapsulating Carrot Somatic Embryo [0057] Carrot cultured cells and somatic embryos were produced as follows.
"Plant cell tissue culture" using carrot roots (1979
Year, Harada, Komamine, Rigaku Kogyo) p. 91-104. From the somatic embryos produced in the liquid medium, somatic embryos having a size of 1.4 mm to 1.7 mm were selectively obtained using a mesh of 1.4 mm and a 1.7 mm nylon mesh. These selected somatic embryos had shapes ranging from heart-shaped to torpedo-shaped. These adventitious embryos were suspended in an MS medium containing no hormone and containing 0.2% of chitosan to obtain the contents of the capsule.

In the encapsulation, the contents are changed, and the pump is adjusted so that the adventitious embryos of the contents liquid (the innermost layer) enter one by one.
The procedure was carried out in the same manner as in Example 1 except that the lipase to be added to the gelatin of the outer coat layer was 0.05% by a submerged dropping method using a sterile seamless soft capsule manufacturing machine with a triple tube nozzle. The particle size of the obtained artificial seed of the encapsulated somatic embryo in which the outermost layer was dried was 7 mm.

As Comparative Example 1, encapsulated carrot somatic embryos were prepared in the same manner as in Example 2 except that lipase was not added to the gelatin film layer.

As Comparative Example 2, a carrot somatic embryo embedded with alginate gel beads was prepared. That is, the somatic embryos produced and selected in Example 2 were suspended in an MS medium containing 3% (w / v) sodium alginate, and added dropwise to a 50 mM calcium chloride solution, thereby embedding the somatic embryos in calcium alginate gel beads. (Particle size: about 6 mm) was prepared. This is the same as that conventionally reported as an artificial seed.

(Sprouting test 1: Effect of adding lipase to outer shell layer) An artificial seed containing lipase in the outer shell layer in which the carrot somatic embryo was encapsulated (Example 2) and an artificial seed not containing lipase (Comparative Example) 1) was prepared. Immediately after the preparation, 100 seeds of each artificial seed were seeded at a depth of 3 cm from the topsoil into a pot containing soil in a field not subjected to soil sterilization, and watered. After seeding, the seeds were placed in an incubator at 25 ° C. and watered once every two days while illuminating at 3000 Lux for 16 hours per day. One week later, two weeks later,
Table 2 shows the results of counting the number of germinations after 3 weeks.

[0062]

[Table 2]

As shown in Table 2, the artificial seeds containing lipase in the outer coat layer rapidly decomposed the hardened oil and fat in the inner coat layer,
Germination was observed in two weeks, whereas lipase-free artificial seeds did not germinate in two weeks, indicating that germination took three weeks. From these results, it is shown that the capsule of the present invention in which the disintegration is controlled is superior as an artificial seed because the inner skin layer (hardened oil and fat) collapses and germinates much more quickly than the conventional capsule. Was.

(Preservation and Germination Test 2) The artificial seed of the capsule-embedded somatic embryo containing lipase in the outer coat layer prepared in Example 2 and the somatic embryo embedded in calcium alginate gel beads of Comparative Example 2 were treated at 20 ° C. 2 in a 50% humidity incubator
Stored for months. Although no change in appearance was observed in the encapsulated artificial seed of Example 2, the calcium alginate gel beads of Comparative Example 2 were considerably dried on the third day, the beads became smaller, and the adventitious embryos appeared in one week. It was dry and dry. At the time of preparing the artificial seeds, one month and two months later, 100 seeds each were seeded at a depth of 2 cm from the topsoil into a pot filled with soil from a field not subjected to soil sterilization, and watered. After sowing, the pot was cooled at 25 ° C and 3000 Lu
x for 16 hours in an incubator illuminated, watered once every two days, and germinated after 2 weeks each. Table 3 shows the results.

[0065]

[Table 3]

As shown in Table 3, the germination rate immediately after preparation was almost the same in both Example 2 and Comparative Example 2. However, when stored for one month or more, Comparative Example 2 which is a conventional type artificial seed was obtained. Did not germinate at all. Germination was observed at a high rate in the soft encapsulated artificial seeds of the present invention. From these results, it was shown that the soft capsule of the present invention containing an adventitious embryo is superior to the conventional adventitious seed of the adventitious embryo type embedded with gel alginate in terms of storage stability as an artificial seed. Also, since it germinates at a high rate even after long-term storage,
It was shown to be practical as an artificial seed.

Example 3 Artificial Seeds Encapsulating Strawberry Leaf Buds A shoot apex tissue extracted from cultivated strawberries is placed on an MS medium containing 10 g / l of sucrose and 2 g / l of gellan gum as a gelling agent. After culturing at 25 ° C. in the dark for 14 days, the cells were transferred to an MS liquid medium containing 0.2 mg / l of benzyladenine and 10 g / l of sucrose,
Rotational shaking culture was performed at 150 rpm at 25 ° C. while irradiating at 2000 Lux for 16 hours per day. 3
After culturing for 0 days, the obtained leaf buds were
using a nylon mesh of 1.4 mm,
Leaf buds of 2 mm size were obtained. The obtained leaf buds were suspended in a hormone-free MS liquid medium and used as the contents of the capsule.

For encapsulation, the contents were changed to a leaf bud suspension, the pump was adjusted so that the leaf buds of the content liquid (innermost layer) entered one by one, and lipase added to gelatin in the outer coat layer was 0.05%. The procedure was performed in the same manner as in Example 1 except that the method was as described in Example 1, using a triple tube nozzle aseptic seamless soft capsule manufacturing machine by a submerged dropping method. The obtained capsules were also dried at a low temperature in the same manner as in Example 1 to obtain artificial seeds of dried and encapsulated strawberry leaf buds. The particle size after drying the outermost layer is 8 mm
Met.

On the other hand, as Comparative Example 3, an encapsulated strawberry leaf bud having a dried outer coat layer was prepared in the same manner as in Example 3 except that lipase was not added to the gelatin solution.

(Sprouting Test 3) As in the case of the germination test in Example 2, encapsulated strawberry leaf-bud artificial seeds containing lipase in the outer coat layer of Example 3 and encapsulated strawberries not containing lipase of Comparative Example 3 20 artificial leaf seeds
It was stored in an incubator at 50 ° C. and a humidity of 50% for 2 months, and a germination test was performed in the same manner as in the germination test 1. Table 4 shows the results
Shown in

[0071]

[Table 4]

As shown in Table 4, even in artificial seeds of encapsulated strawberry leaf buds, the germination of Example 3 in which lipase was added to the capsule outer layer was faster than that of Comparative Example 3 in which lipase was not added. It has been shown.

Further, even after storage for 2 months, the artificial seeds of encapsulated strawberry leaf buds in which lipase was added to the outer coat layer showed rapid germination at a high rate, and were more effective than those in which no lipase was added. It was shown to be much better.

Example 4 Pharmaceutical Capsules Cilostazol having a platelet aggregation inhibitory action was suspended in a hardened fat and oil having a melting point of 42 ° C. at a ratio of 1 g / 3 g (w / w). ), And only the hardened oil and fat containing nothing was used as the inner film layer. As an outer coat layer, pectin is 5% (w
/ W) Gelatin 20% (w / v) solution containing a powdery lipase for food (Lilipase A-10FG, manufactured by Nagase Seikagaku Corporation) added at a rate of 0.05% (w / w). Using. These are encapsulated in coagulated oil using a triple tube nozzle seamless soft capsule manufacturing machine,
After deoiling immediately, the drum was dried while sending dry air at 10 ° C. The particle size of the dried capsule was 2.5 mm. 1 g of this capsule contains 100 m of cilostazol
g is included.

Separately, as Comparative Example 4, a capsule having a dried outer skin layer was prepared in the same manner as in Example 5 except that lipase was not added to the outer skin layer.

(Absorption test) As a group of 6 adult female big dogs, 1 g of a capsule containing lipase was orally administered to the outer coat layer prepared in Example 4 respectively. As a control, 1 g of a capsule containing no lipase in the outer coat layer prepared in Comparative Example 4 was orally administered to another group of six animals.
Blood was sampled over time, the time required for cilostazol to reach the maximum blood concentration was determined, and the average value of 6 animals in each group was compared. As a result, the average value of the group using the capsule of Comparative Example 4 was 6 hours, whereas the average value of the group using the capsule of Example 4 was as short as 4.5 hours. That is, it was found that the capsule in which the lipase was added to the outer coat layer of Example 5 collapsed the inner coat layer in a shorter time than Comparative Example 4, and the drug was absorbed from the intestine. Since the medicament capsule of the present invention is expected to promptly show the effect of the drug contained therein, the superiority of the capsule of the present invention is apparent.

Example 5 Capsules Containing Microorganisms in Outer Coating Layer Candida rugosa IFO 0591, which is known to secrete lipase,
O No. Cultured in 108 liquid medium, 100 ml of culture solution
Was centrifuged at 8000 rpm for 20 minutes at 4 ° C., and the supernatant was discarded to collect the cells, and the cells were resuspended in 100 ml of water. This suspension was added to a 10% (v / w) solution in a 22% (w / w) gelatin solution.
v) was added and mixed to obtain a solution for the outer coating layer. Otherwise, the red pigment was encapsulated in the same manner as in Example 1, dried, and further dried in a desiccator containing concentrated sulfuric acid.
Dried for hours.

These capsules were stable after being stored in a polyethylene bag containing silica gel and stored at 20 ° C. for 3 months without the red color of the content liquid appearing on the surface.

The capsules immediately after preparation and the capsules stored for 3 months under the above-mentioned conditions were placed in a Petri dish with 50 tablets each, sprayed with water uniformly by spraying, and kept warm in a 25 ° C. incubator. The time during which red spots of the content liquid were observed on the surface was observed over time, and 50 particles were averaged to determine the film disintegration time. Table 5 shows the results.

[0080]

[Table 5]

As shown in Table 5, in the capsule containing the lipase in the outer coat layer, the disintegration time of the coat was remarkably shortened. Although the capsule of the present invention is a kind of dried and immobilized cells, it is stable after storage for 3 months at room temperature in a dry state, the disintegration time is not so large as about 6%, and it can withstand long-term storage. It has been shown.

[0082]

The capsule of the present invention in which the enzyme or microorganisms that decompose the film layer is contained in the film layer decomposes the film layer according to the activity of the enzyme when water is given, so that the desired time can be obtained. The film layer can be disintegrated at the place, allowing quick release of the contents, contact of the contents with the outside, mixing, artificial seeds, drugs for drug delivery systems,
It can be applied to sequestration of unstable components and sustained release pesticides.

[Brief description of the drawings]

FIG. 1 is a schematic view showing the shape of an artificial seed of the present invention.

──────────────────────────────────────────────────の Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) // A61K 31/4709 A61K 31/4709 (72) Inventor Yoshimasa Kamaguchi 1-chome, Tamazuki, Chuo-ku, Osaka-shi, Osaka 1-30 No. 1 in Morishita Jintan Co., Ltd. (72) Inventor Yuzo Kawahara 1-30 No. 1 Tamazo, Chuo-ku, Osaka City, Osaka Prefecture (72) Inventor Hideki Sunohara 1-chome, Tamazuki, Chuo-ku, Osaka City, Osaka Prefecture No. 1-30 Morishita Jintan Co., Ltd. F term (reference) 2B030 AA02 AB03 CD02 CD03 CD07 CD09 CD14 CD15 CD21 CD24 4C076 AA56 AA57 BB01 CC11 4C086 AA01 BC62 GA07 MA01 MA04 MA37 MA52 NA11 ZA54

Claims (8)

[Claims] A soft capsule having a multilayer structure having an innermost layer and one or more coating layers sequentially laminated so as to cover the innermost layer, wherein at least one of the coating layers contains an enzyme or a microorganism. A soft capsule comprising, wherein the enzyme or the microorganism decomposes a film layer or another film layer containing the enzyme under predetermined conditions. 2. The soft capsule according to claim 1, wherein the enzyme or the microorganism is contained in an outermost coating layer of the soft capsule, and the outermost coating layer is dried. 3. The capsule according to claim 3, wherein the soft capsule is a three-layer capsule having an innermost layer, an inner coating layer covering the innermost layer, and an outer coating layer covering the inner coating layer. The soft capsule according to claim 1, wherein the soft capsule contains an enzyme or a microorganism capable of decomposing the skin layer. 4. The soft capsule according to claim 1, wherein the coating layer is formed of a substance selected from the group consisting of proteins, polysaccharides, hardened fats and oils, and biodegradable plastics. The soft capsule according to the item. 5. The soft capsule according to claim 1, wherein the enzyme is a protease, a polysaccharide-degrading enzyme, a hardened fat / oil-degrading enzyme, or an esterase. 6. The innermost layer contains a medicament, a pesticide or a regenerable plant cell tissue, the inner coat layer is mainly composed of hardened oil and fat, and the outer coat layer is capable of decomposing hardened oil and fat. The soft capsule according to claim 3, which is a dried outer skin layer containing the obtained enzyme or microorganism. 7. The soft capsule according to claim 1, wherein the soft capsule is selected from the group consisting of a seamless soft capsule and a seam soft capsule. 8. An artificial seed comprising a three-layer soft capsule having an innermost layer containing a regenerated plant cell tissue, an inner coat layer covering the innermost layer, and an outer coat layer covering the inner coat layer. An artificial seed, wherein the inner skin layer is mainly composed of hardened fat and the outer skin layer is a dried outer skin layer containing an enzyme or a microorganism capable of decomposing the hardened fat.